Intermolecular cleavage by UmuD-like mutagenesis proteins.
The activity of a number of proteins is regulated by self-processing reactions. Elegant examples are the cleavage of the prokaryotic LexA and lambdaCI transcriptional repressors and the UmuD-like mutagenesis proteins. Various studies support the hypothesis that LexA and lambdaCI cleavage reactions are predominantly intramolecular in nature. The recently described crystal structure of the Escherichia coli UmuD' protein (the posttranslational cleavage product of the UmuD protein) suggests, however, that the region of the protein corresponding to the cleavage site is at least 50 A away from the catalytic active site. We considered the possibility, therefore, that the UmuD-like proteins might undergo self-processing that, in contrast to LexA and lambdaCI, occurs via an intermolecular rather than intramolecular reaction. To test this hypothesis, we introduced into E. coli compatible plasmids with mutations at either the cleavage or the catalytic site of three UmuD-like proteins. Cleavage of these proteins only occurs in the presence of both plasmids, indicating that the reaction is indeed intermolecular in nature. Furthermore, this intermolecular reaction is completely dependent upon the multifunctional RecA protein and leads to the restoration of cellular mutagenesis in nonmutable E. coli strains. Intermolecular cleavage of a biotinylated UmuD active site mutant was also observed in vitro in the presence of the wild-type UmuD' protein, indicating that in addition to the intact UmuD protein, the normal cleavage product (UmuD') can also act as a classical enzyme.
McDonald
J P
JP
Section on DNA Replication, Repair, and Mutagenesis, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-2725, USA.
Frank
E G
EG
Levine
A S
AS
Woodgate
R
R
D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D004259:DNA-Directed DNA Polymerase; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins; D016296:Mutagenesis; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011938:Rec A Recombinases / Q000378:metabolism
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D011938:Rec A Recombinases; D004259:DNA-Directed DNA Polymerase; C055218:UmuD protein, E coli